摘要
采用RT-PCR和RACE技术扩增了杜氏盐藻小G蛋白基因cDNA全长序列(GenBank Accession No.JN989548),命名为DsRab,对其进行生物信息学分析,并通过实时荧光定量PCR方法检测盐胁迫下该基因的表达情况。结果表明,DsRab基因的cDNA全长为1 299 bp,开放阅读框(ORF)为612 bp,编码203个氨基酸,5'非编码区78 bp,3'非编码区609 bp;保守性结构域分析可知编码的小G蛋白有4个GTP/GDP保守结构域,1个效应区、1个羧基端的半胱氨酸结构域和5个Rab亚家族共有的结构域;二级结构预测表明该蛋白有32.02%的α-螺旋,23.65%的伸展片段,44.33%的自由卷曲,三维建模成功;比对分析发现DsRab蛋白与多种生物的Ypt/Rab的氨基酸序列具有较高的同源性。荧光定量PCR结果表明,盐藻在高盐(3.0 mol/L)胁迫下,DsRab基因表达量显著上调,1 h后表达量达到最大值,为正常培养下对照组(0 h)的4.9倍,差异极显著(P<0.01)。
Dunaliella salinasmall GTP-binding Protein Gene（GenBank Accession No.JN989548）was cloned by RT-PCR and RACE technology,namedDsRab,the bioinformatics-analysis was performed,and the DsRabgene expression pattern was studied under 3 mol/L NaCl by Real-time quantitative RT-PCR method.The results showed that the full length of cDNA for DsRabGene was 1 299 bp,which contains 78 bp 5＇UTR,609 bp 3＇UTR and 612 bp ORF.The ORF codes a 203 amino acids protein with four GTP/GDP binding conserved domains,one effector domain,a cysteine residues at the COOH-terminus,and five common domain of Rab sub-family;Secondary structure prediction indicated that the α-helix,β-strands and random coil in it were 32.02% of,23.65% and 44.33%.Protein homologue analysis showed that the DsRab shared high homology with Ypt/Rab from other species.The real-time quantitative PCR（RT-PCR）revealed that the expression of DsRabgene was increased by 4.9-fold under 3 mol/L NaCl comparising with that under normal level（including 1.5 mol/L NaCl）（P0.01）.
出处
《生物技术通报》
CAS
CSCD
北大核心
2013年第9期77-83,共7页
Biotechnology Bulletin
基金
国家自然科学基金项目(30972240)
辽宁省教育厅科技研究项目(2008T023)
